Macrolide antibiotics, characterized by a large lactone ring to which are attached one or more deoxy sugars, usually cladinose and desosamine, are antimicrobial drugs that are active against aerobic and anaerobic gram positive cocci and are prescribed for the treatment of respiratory tract and soft tissue infections. The macrolides, which belong to the polyketide class of natural products, function by reversibly binding to the 23S rRNA component of the 505 subunit of the bacterial ribosome, blocking protein synthesis and preventing bacterial growth and reproduction. The macrolide antibiotics bind to the large ribosomal subunit and inhibit protein synthesis by blocking the path of the nascent peptide in the exit tunnel (Gaynor, M., and A. S. Mankin. 2003. Macrolide antibiotics: binding site, mechanism of action, resistance. Curr. Top. Med. Chem. 3:949-961). The chemical structure of the prototype macrolide erythromycin A is represented by a 14-atom lactone ring substituted with 3-O-cladinose and 5-O-desosamine sugar residues. Macrolides of the subsequent generations differ in the structures of the lactone, such as azithromycin, as well as the number, composition, and sites of attachment of the side chains (Franceschi, F., Z. Kanyo, E. C. Sherer, and J. Sutcliffe. 2004. Macrolide resistance from the ribosome perspective. Curr. Drug Targets Infect. Disord. 4:177-191; Sutcliffe, J. A. 2005. Improving on nature: antibiotics that target the ribosome. Curr. Opin. Microbiol. 8:534-542).
The binding site of macrolides in the ribosome includes the 23S rRNA residues A2058, A2059, A2062, A2503, G2505, and C2611 [or U2611] (using here and throughout the E. coli numbering, see Tu, D., G. Blaha, P. B. Moore, and T. A. Steitz. 2005. Structures of MLSBK antibiotics bound to mutated large ribosomal subunits provide a structural explanation for resistance. Cell 121:257-270). It is to be understood that the corresponding nucleic acid residues in ribosomes of other organisms are described herein by reference to the E. coli nucleic acids. One of the main mechanisms of resistance to macrolide antibiotics is based on dimethylation of A2058 by methyltransferase encoded in erm genes (Weisblum, B. 1995. Erythromycin resistance by ribosome modification. Antimicrob. Agents Chemother. 39:577-585). Erm-catalyzed dimethylation of A2058 leads to a steric clash with the drug and reduces affinity of erythromycin for the ribosome. Similar to several other antibiotic resistance genes, erm genes are often inducible by erythromycin and similar drugs. In an effort to combat resistance, a newer class of macrolides, known as ketolides, was developed (Bryskier, A. 2000. Ketolides-telithromycin, an example of a new class of antibacterial agents. Clin. Microbiol. Infect. 6:661-669). Ketolides show improved activity against strains with inducible erm genes and are believed to exhibit a tighter binding to the ribosome compared with macrolides of the previous generations (Poehlsgaard, J., and S. Douthwaite. 2003. Macrolide antibiotic interaction and resistance on the bacterial ribosome. Curr. Opin. Investig. Drugs 4:140-148; Poehlsgaard, J., P. Pfister, E. C. Bottger, and S. Douthwaite. 2005. Molecular mechanisms by which rRNA mutations confer resistance to clindamycin. Antimicrob. Agents Chemother. 49:1553-1555). In ketolides, the 3-O-cladinose is replaced by a keto function (hence the name of the class); a cyclic carbamate is fused at the C11-C12 position; and an extended side chain, such as an alkyl side chain bearing a an aryl or heteroaryl group, which may be substituted, is attached at the C11-N atom (11-N) of the carbamate, as in the ketolide telithromycin, or at another position of the lactone ring, such as the 6-O— position as in cethromycin. Early biochemical and genetic studies showed that the extended side chain of ketolides establishes important new interactions with the ribosome that might account for increased efficacy of these drugs. Specifically, chemical probing and resistance mutations pointed to interactions of the 11-N-side chain of telithromycin with the rRNA residues in the loop of helix 35 of the E. coli 23S rRNA and with U2609 (Garza-Ramos, G., L. Xiong, P. Zhong, and A. Mankin. 2002. Binding site of macrolide antibiotics on the ribosome: new resistance mutation identifies a specific interaction of ketolides with rRNA. J. Bacteriol. 183:6898-6907; Hansen, L. H., P. Mauvais, and S. Douthwaite. 1999. The macrolide-ketolide antibiotic binding site is formed by structures in domains II and V of 23S ribosomal RNA. Mol. Microbiol. 31:623-632; Xiong, L., S. Shah, P. Mauvais, and A. S. Mankin. 1999. A ketolide resistance mutation in domain II of 23S rRNA reveals proximity of hairpin 35 to the peptidyl transferase centre. Mol. Microbiol. 31:633-639 46). However, subsequent crystallographic studies of the first clinically approved ketolide telithromycin bound to the bacterial (Deinococcus radiodurans) or archaeal (Haloarcula marismortui) ribosome showed the placement of the 11N-side chain in a position that appeared incompatible with rRNA protections and mutations observed in the E. coli ribosome (Berisio, R., J. Harms, F. Schluenzen, R. Zarivach, H. A. Hansen, P. Fucini, and A. Yonath. 2003. Structural insight into the antibiotic action of telithromycin against resistant mutants. J. Bacteriol. 185:4276-4279; 41). Furthermore, orientation of the 11-N-side chain differed significantly between the reported D. radiodurans and H. marismortui structures, therefore leaving open the question of how telithromycin would bind to the ribosomes of bacteria targeted by ketolide antibiotics, including Gram positive pathogenic bacteria.
Although telithromycin, the first ketolide introduced into medical practice in 2001, showed excellent activity against many strains of Gram positive pathogens, the safety issues that became apparent upon the wider use of the drug have curbed its clinical use (reviewed in Rafie, S., C. MacDougall, and C. L. James. 2010. Cethromycin: a promising new ketolide antibiotic for respiratory infections. Pharmacotherapy 30:290-303). The adverse effects associated with telithromycin spurred a search for newer ketolides. One of the novel promising drugs of this class is the fluoroketolide CEM-101 disclosed in international patent application, publication number WO 2004/080391, and its counterpart publication US 2006/0100164, at Example 7 and identified as OP-1068. The structure of CEM-101 is similar to that of telithromycin except for the presence of a fluorine atom at C2 of the lactone and the differing aromatic groups on the 11-N side chain (which in CEM-101 is a 4-[4-(3-aminophenyl)[1,2,3]triazol-1-yl]butyl group). Additional fluoroketolides are reported in C.-H. Liang et al., Bioorg. Med. Chem. Lett. 15 (2005) 1307-1310.
In microbiological tests involving a number of clinical pathogens, CEM-101 is characterized by lower minimal inhibitory concentration (MIC) values compared with telithromycin and exhibits enhanced activity against resistant organisms, including telithromycin-intermediate and telithromycin-resistant organisms (McGhee, P., C. Clark, K. M. Kosowska-Shick, K. Nagai, B. Dewasse, L. Beachel, and P. C. Appelbaum. 2010. In vitro activity of CEM-101 against Streptococcus pneumoniae and Streptococcus pyogenes with defined macrolide resistance mechanisms. Antimicrob. Agents. Chemother. 54:230-238). Furthermore, in comparison with telithromycin and cladinose-containing macrolides CEM-101 shows significantly enhanced accumulation in the macrophages.
It has been discovered that upon analysis of the binding interactions of CEM-101 compared to telithromycin and other macrolides, ketolides may be designed with high activity against pathogenic bacteria and against resistant strains of bacteria, including strains resistant to other ketolides like telithromycin.
In one illustrative embodiment of the invention, compounds of formula (I)A-L-Q  (I)are described herein, and pharmaceutically acceptable salts, hydrates, solvates, and prodrugs thereof, wherein:
A is a moiety capable of forming one or more of the hydrogen bond interactions with one or more of A752, G748, and G748;
L is a linking chain consisting of one or more divalent radicals selected from the group consisting of amino, O, and S, and alkylene, heteroalkylene, cycloalkylene, heterocycloalkylene, alkenylene, heteroalkenylene, cycloalkenylene, heterocycloalkenylene, arylene, and heteroarylene, each of which is optionally substituted, provided that L does not include an 0-0 or O—S; and
Q is a 3-keto- or 2-fluoro-3-keto macrolactone capable of binding in the major macrolide site in the upper part of a bacterial ribosomal exit tunnel.
In another embodiment, compounds of formula (I) are described where Q is of the formula
in which L is bonded to the 11-N of Q;
R2 is H or F;
R5 is an aminosaccharide residue; and
R6 is H or (1-6C)alkyl; and
wherein A-L- is other than 4-[4-(3-aminophenyl)[1,2,3]triazol-1-yl]butyl or 4-[4-(6-aminopyridin-2-yl)[1,2,3]triazol-1-yl]butyl bonded to the 11-N of Q.
In another embodiment, compounds of formula (I) are described herein, and pharmaceutically acceptable salts, hydrates, solvates, and prodrugs thereof, wherein (a) A is a moiety capable of forming one or more of the hydrogen bond interactions with one or more of O4′ of A752 (as a donor), O6 of G748 (as a donor), and N1 of G748 (as an acceptor). In another embodiment, (b) the 3-keto group of Q is capable of forming a hydrogen bond interaction with U2609. In another embodiment, (c) the aminosaccharide of Q is capable of forming a hydrogen bond interaction with A2059. In another embodiment, (d) the aminosaccharide of Q is capable of forming a hydrogen bond interaction with G2505. In another embodiment, compounds of formula (I) are described herein, and pharmaceutically acceptable salts, hydrates, solvates, and prodrugs thereof, wherein the compound exhibits any combination of (a), (b), (c), and (d).
It is to be understood that the hydrogen bond forming capability of A may be determined using any conventional method. For example, hydrogen bond forming capability of A may be determined by computer modeling the compound in the E. coli 23S ribosomal site; or by co-crystallizing the compound in the E. coli 23S ribosomal site.
It is to be further understood that the hydrogen bond forming capability of the 3-keto group of Q may be determined using any conventional method. For example, in another embodiment, the compound protects U2609 from modification by CMCT, such as protecting against modification to a greater extent than does telithromycin, in RNA footprinting.
It is to be further understood that the hydrogen bond forming capability of the aminosaccharide of Q may be determined using any conventional method. For example, in another embodiment, the compound protects G2505 from modification by kethoxal, such as protecting against modification to a greater extent than does erythromycin, clarithromycin, azithromycin, and/or telithromycin, in RNA footprinting. In another embodiment, the compound blocks methylation of A2059 by DMS, such as blocking methylation to a greater extent than does erythromycin, clarithromycin, azithromycin, and/or telithromycin, in RNA footprinting.
In alternatives of each of the embodiments described herein, the bacteria may be a resistant strain. Illustrative resistant strains, include but are not limited to erythromycin resistant strains, clarithromycin resistant strains, azithromycin resistant strains, telithromycin resistant strains, mefA resistant strains, and ermB resistant strains.